Fig. 5.

Mouse lung chronically depleted of ITSN-1s show phenotypically-altered ECs. A BrdU/FITC Ab immunostaining (a1, a4, a5), followed by CD31/AlexaFluor594 (a2, a6, a7) shows BrdU positive ECs within alveolar wall, at 10d (a3, arrows) and 24d (a8, a9, arrows). Arrowheads (a8, a9) point towards BrdU positive alveolar epithelial cells. Bars: 20 μm, (a1–a9). B Representative Western blots of lung lysates of control, wt-mice (a), siRNActrl (b) and veh (c), 72 h post-injection as well as siRNA_ITSN-treated mice, at the time points indicated, using phospho-specific Erk1/2^MAPK and total Erk1/2^MAPK Abs. The graph shows densitometric analysis of Erk1/2^MAPK activation (phospho-Erk1/2 relative to total Erk1/2) as mean ± SEM of 3 separate experiments. C Expression TGFβ, BMP-2/4 and VEGF-A, in mouse lung lysates of control and siRNA_ITSN-treated mice, at the time points indicated assessed by Western blot with specific anti-TGFβ, BMP-2/4 and VEGF-A Abs. Actin was used as loading control. D. Densitometric analysis of TGF-β, BMP-2/4 and VEGF-A immunoreactivity. Different Abs and detection conditions did not allow a quantitative assessment of the ratio between the growth factors. E. Evaluation of Bad phosphorylation by Western blotting with anti-phospho Ser¹¹²-Bad, Ser¹³⁶-Bad and Ser¹⁵⁵-Bad Abs. The blots were stripped and reprobed with an anti-Bad Ab. The Western blots C, E are representative for 3–4 different experiments; densitometric analyses B, D were applied on 3 different HyBlot CL films. Densitometric values ± SEM are representative for 3 independent experiments